Combined immunofluorescence, RNA fluorescent in situ hybridization, and DNA fluorescent in situ hybridization to study chromatin changes, transcriptional activity, nuclear organization, and X-chromosome inactivation.
Epigenetic mechanisms lead to the stable regulation of gene expression without alteration of DNA and trigger initiation and/or maintenance of cell-type-specific transcriptional profiles. Indeed, modulation of chromatin structure and the global 3D organization of the genome and nuclear architecture participate in the precise control of transcription. Thus, dissection of these epigenetic mechanisms is essential for our understanding of gene regulation. In this chapter, we describe challenging combinations of immunofluorescence, and RNA and DNA fluorescent in situ hybridization and their application to our studies of a remarkable example of epigenetic control of gene expression in female mammals, the process of X chromosome inactivation.
Chaumeil
Julie
J
Mammalian Developmental Epigenetics Group, CNRS UMR 218, Curie Institute, Paris, France.
Augui
Sandrine
S
Chow
Jennifer C
JC
Heard
Edith
E
D000818:Animals; D002467:Cell Nucleus / Q000235:genetics; D002843:Chromatin / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D053595:Embryonic Stem Cells; D044127:Epigenesis, Genetic; D005260:Female; D005786:Gene Expression Regulation; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D012313:RNA / Q000378:metabolism*; D014158:Transcription, Genetic; D014960:X Chromosome / Q000378:metabolism; D049951:X Chromosome Inactivation
D002843:Chromatin; D012313:RNA; D004247:DNA